OBJECTIVE: To define the sites of Mamu-AG mRNA expression during fetal and placental development in the rhesus monkey. RESULTS Previous studies demonstrated that the rhesus monkey placenta expresses a mRNA for a unique MHC class I molecule which has been designated Mamu-AG, which may be a functional homolog of HLA-G that is expressed in the human placenta. MHC class I immunoactivity localized to syncytiotrophoblast (ST), but not in villous cytotrophoblast (CT). Low levels of patchy staining were seen with extravillous trophoblast (ET). This contrasts with ET staining of HLA-G in the human. Allele-specific in situ hybridization showed a pattern of mRNA expression identical to immunostaining, indicating Mamu-AG gene transcription specifically in ST. The question arises as to whether Mamu-AG expression is restricted to the placenta, or whether it may be expressed in other fetal or adult tissues, as has been reported for HLA-G. Primers which hybridized with the divergent alpha-1 domain were designed for Mamu-AG-specific PCR amplification. Using these primers, the expression of Mamu-AG mRNA was detectable in a number of tissues, most consistently in the pituitary gland and the kidney. Since PCR is highly sensitive but generally also highly qualitative, we also developed a Mamu-AG-specific ribonuclease protection assay to compare relative Mamu-AG mRNA levels in these tissues. The expression of Mamu-AG was abundant in the placenta, very low in the pituitary gland, and undetectable in other nonplacental tissues. FUTURE DIRECTIONS This information is important in developing in vivo approaches to examine Mamu-AG function. We plan to develop monoclonal antibodies to Mamu-AG to confirm very low mRNA expression in nonplacental tissues, and to interface with Mamu-AG function in vivo and in vitro. KEY WORDS placenta, major histocompatibility complex, maternal-fetal immune tolerance